Apoptosis, or programmed cell death in multicellular organisms, is one of the fundamental means by which a cell can respond to environmental changes. One of the best studied mammalian apoptosis systems involves Fas (also designated APO-1 and CD95), a type I membrane receptor that, when crosslinked by its cognate ligand, induces apoptosis in a wide variety of cells (for review, see Nagata, 1994). The extracellular interaction of Fas ligand with the cell membrane-spanning Fas receptor activates an intracellular signal transduction cascade finally activating proteases in the IL-1.beta.-converting enzyme (ICE) family (Henkart, 1996). Transduction of an apoptosis signal depends on interaction between the intracellular "death domain" of Fas with a cytoplasmic 23-kDa protein, MORT1 [(Boldin, et al., 1995), also termed FADD (Chinnaiyan, et al., 1995)]. The events leading from the production of an activated Fas trimer complex to cell destruction mediated by ICE-like proteases are yet to be determined, but recruitment of two MORT1/FADD molecules into a death-inducing signaling complex with the death domain of Fas appears to be a necessary step (Kischkel, et al., 1995). The end result of this pathway is cell death by a distinctive mechanism characterized by nuclear and cytoplasmic condensation and DNA fragmentation.
The human MORT1/FADD gene spans approximately 3.6 kb and contains two exons (286 and 341 bp) separated by a 2.0-kb intron. MORT1/FADD was mapped to chromosome 11q13.3 by the independent techniques of PCR screening of somatic cell hybrid mapping panels and fluorescence in situ hybridization (Kim, et al., 1996). Knowledge of the chromosomal location and gene structure of MORT1/FADD will assist efforts to determine its involvement in genetic disorders of apoptosis. Defects in apoptosis due to mutations in the Fas receptor have been described (Fisher, et al., 1995; Rieux-Laucat, et al., 1995) in patients with a rare autoimmune lymphoproliferative syndrome (ALPS) including nonmalignant lymphadenopathy, hepatosplenomegaly, and expanded populations of CD3.sup.+ CD4.sup.- CD8.sup.- lymphocytes. However, the existence of some patients with clinical findings of ALPS, but no Fas mutation, suggests that defects in other proteins in the Fas pathway may also produce ALPS. The functional involvement of MORT1/FADD in the Fas pathway makes it a candidate for mutation analysis in ALPS.
Localization of MORT1/FADD to 11q13.3 also makes it a candidate for human diseases associated with this part of the genome. IDDM4, a predisposing locus for familial insulin-dependent diabetes mellitus (IDDM) has been mapped to this region by linkage studies (Cordell, et al., 1995; Davies, et al., 1994; Hashimoto, et al., 1995). The pathogenesis of IDDM may involve autoimmune T lymphocyte-mediated destruction of pancreatic .beta. islet cells that produce insulin (Tisch and McDevitt, 1996). MORT1/FADD's chromosomal location, coupled with its known role in lymphocyte apoptosis, makes it a candidate for mutational analysis in patients with IDDM4-linked familial diabetes.
In addition, the 11q13 region is amplified in several human malignancies, including carcinoma of the breast, bladder, esophagus, head and neck, and lung (Schuuring, 1995; Szepetowski, et al., 1995). Amplification of this region has been associated with poor prognosis in patients with operable breast cancer (Schuuring, et al., 1992). The mapping of MORT1/FADD to the amplicon of the breast cancer cell line MDA-MB-134-VI (Lafage, et al., 1992) raises the possibility of its involvement in tumor growth. Future linkage and mutation analysis of MORT1/FADD in other diseases may support the hypothesis that disregulation of cell death is a fundamental mechanism for the pathogenesis of human disease (Thompson, 1995).
A single gene can encode more than one mRNA transcript by transcriptional processing events such as use of alternative promoters, alternative splicing, and alternative polyadenylation (Farrow, 1997; Lewin, 1994). The resulting variant transcript isoforms may differ in stability, translatability, or protein sequence encoded, each of which may impact upon the function of the encoded protein. Transcript variants resulting from alternative RNA processing can be tissue-specific, developmentally regulated, endocrine regulated, or can appear in response to specific exogenous cues. In addition, transcript variants of a gene can result from differences in genomic sequence (among individuals or among cell lines).
To date, only one transcript isoform has been reported for the MORT1/FADD gene (Boldin, et al., 1995; Chinnaiyan, et al., 1995). This invention describes the identification of two new isoforms of MORT1.